Hyphal Wall Research Articles

Candida krusei and Candida glabrata reduce the filamentation of Candida albicans by downregulating expression of HWP1 gene.

Pathogenicity of Candida albicans is associated with its capacity switch from yeast-like to hyphal growth. The hyphal form is capable to penetrate the epithelial surfaces and to damage the host tissues. Therefore, many investigations have focused on mechanisms that control the morphological transitions of C. albicans. Recently, certain studies have showed that non-albicans Candida species can reduce the capacity of C. albicans to form biofilms and to develop candidiasis in animal models. Then, the objective of this study was to evaluate the effects of Candida krusei and Candida glabrata on the morphogenesis of C. albicans. Firstly, the capacity of reference and clinical strains of C. albicans in forming hyphae was tested in vitro. After that, the expression of HWP1 (hyphal wall protein 1) gene was determined by quantitative real-time PCR (polymerase chain reaction) assay. For both reference and clinical strains, a significant inhibition of the hyphae formation was observed when C. albicans was incubated in the presence of C. krusei or C. glabrata compared to the control group composed only by C. albicans. In addition, the culture mixed of C. albicans-C. krusei or C. albicans-C. glabrata reduced significantly the expression of HWP1 gene of C. albicans in relation to single cultures of this specie. In both filamentation and gene expression assays, C. krusei showed the higher inhibitory activity on the morphogenesis of C. albicans compared to C. glabrata. C. krusei and C. glabrata are capable to reduce the filamentation of C. albicans and consequently decrease the expression of the HWP1 gene.

Alteration in the ultrastructural morphology of mycelial hyphae and the dynamics of transcriptional activity of lytic enzyme genes during basidiomycete morphogenesis.

The morphogenesis of macromycetes is a complex multilevel process resulting in a set of molecular-genetic, physiological-biochemical, and morphological-ultrastructural changes in the cells. When the xylotrophic basidiomycetes Lentinus edodes, Grifola frondosa, and Ganoderma lucidum were grown on wood waste as the substrate, the ultrastructural morphology of the mycelial hyphal cell walls differed considerably between mycelium and morphostructures. As the macromycetes passed from vegetative to generative development, the expression of the tyr1, tyr2, chi1, chi2, exg1, exg2, and exg3 genes was activated. These genes encode enzymes such as tyrosinase, chitinase, and glucanase, which play essential roles in cell wall growth and morphogenesis.

Hyphal wall protein 1 gene: A potential marker for the identification of different Candida species and phylogenetic analysis.

Background and Purpose:Hyphal wall protein 1 (HWP1) is an important adhesin which usually is expressed on the germ tube and hyphal surface produced by different Candida species. The hyphal wall protein-coding gene (HWP1) was evaluated as a novel identification and phylogenetic marker in Candida tropicalis, C. orthopsilosis, C. parapsilosis and C. glabrata.Materials and Methods:Initially, four specific primer pairs were designed, and the target was amplified and finally sequenced. A total of 77 Candida isolates from four different species were included in the study. Consensus sequences were used for the evaluation of phylogenetic tree using the CLC Genome Workbench, GENEIOUS, and MEGA softwares and the levels of nucleotide and amino acid polymorphism were assessed.Results:According to the results, the specific amplified fragments of HWP1 gene were useful for the differentiation of four species. Intra-species variation was observed only in C. tropicalis with two DNA types. The phylogenetic tree of Candida species based on the HWP1 gene showed consistency in topology with those inferred from other gene sequences.Conclusion:We found that HWP1 gene was an excellent marker for the identification of non-albicans Candida species as well as the phylogenetic analysis of the most clinically significant Candida species.

Morphological and phylogenetic studies of two new neotropical species of Loweomyces (Polyporales, Basidiomycota)

The genus Loweomyces is characterized by resupinate to pileate, sessile to stipitate basidiomes, monomitic to dimitic hyphal system with presence of pseudoskeletal hyphae, thin-walled hymenial cystidia, and usually cyanophilous spore and hyphal walls. The genus has not been widely accepted and some authors placed Loweomyces species in Abortiporus, Tyromyces or Antrodiella. During a survey of polypores in southern Brazil, two unidentified species with similar features to Loweomyces spp. were found. Morphological and molecular studies were carried out in order to understand their morphological and evolutionary relationships and to confirm their phylogenetic placement and taxonomic identity. DNA sequences of the internal transcribed spacer and large subunit of ribosomal RNA gene were used for phylogenetic studies. Both studied species have similar morphological characteristics to other Loweomyces spp. and group close to L. fractipes phylogenetically; therefore, Loweomyces spissus and L. tomentosus are proposed as new species. Full descriptions, illustrations of microscopic features, and an identification key to the accepted species in the genus are presented.

Synergic effect of combination of glycyrol and fluconazole against experimental cutaneous candidiasis due to Candida albicans.

In this study, we investigated the anti-fungal activity of glycyrol, a coumarine isolated from licorice (Glycyrrhizae Radix), in a murine model of cutaneous candidiasis caused by Candida albicans. Compared to the infected sites, located on the mice's back, of the untreated control mice, the infected sites treated with glycyrol had reduced CFU (colony forming unit) values up to 60 and 85.5% at 20 and 40μg/mouse of glycyrol, respectively (P<0.01). The antifungal activity of glycyrol was synergistically increased when glycyrol (10μg/mouse) was combined with fluconazole (10μg/mouse), demonstrating that the combination therapy is approximately 4 times more effective than fluconazole alone at 20μg/mouse (P<0.01). Additionally, the combination activity was 1.65 times greater than the antifungal activity of fluconazole alone at 40μg/mouse (P<0.05). In seeking glycyrol's antifungal mechanism, we determined that glycyrol inhibited hyphal induction and cell wall adherence of C. albicans. Thus, it is very likely that, by damaging the cell wall, glycyrol helps fluconazole invade C. albicans more readily and attack fluconazole's target in the fungus membrane. In summary, our data indicate that glycyrol may contribute to the development of a novel agent that possesses antifungal activity against cutaneous candidiasis.

Host-induced silencing of Fusarium culmorum genes protects wheat from infection.

Plants producing antisense or double-stranded RNA molecules that target specific genes of eukaryotic pests or pathogens can become protected from their attack. This beneficial effect was also reported for plant-fungus interactions and is believed to reflect uptake of the RNAs by the fungus via an as yet unknown mechanism, followed by target gene silencing. Here we report that wheat plants pre-infected with Barley stripe mosaic virus (BSMV) strains containing antisense sequences against target genes of the Fusarium head blight (FHB) fungus F. culmorum caused a reduction of corresponding transcript levels in the pathogen and reduced disease symptoms. Stable transgenic wheat plants carrying an RNAi hairpin construct against the β-1, 3-glucan synthase gene FcGls1 of F. culmorum or a triple combination of FcGls1 with two additional, pre-tested target genes also showed enhanced FHB resistance in leaf and spike inoculation assays under greenhouse and near-field conditions, respectively. Microscopic evaluation of F. culmorum development in plants transiently or stably expressing FcGls1 silencing constructs revealed aberrant, swollen fungal hyphae, indicating severe hyphal cell wall defects. The results lead us to propose host-induced gene silencing (HIGS) as a plant protection approach that may also be applicable to highly FHB-susceptible wheat genotypes.

Genomic and functional analyses unveil the response to hyphal wall stress in Candida albicans cells lacking β(1,3)-glucan remodeling.

BackgroundThe cell wall is essential for the yeast to hypha (Y-H) transition that enables Candida albicans to invade human tissues and evade the immune system. The main constituent, β(1,3)-glucan, is remodeled by glucanosyltransferases of the GH72 family. Phr1p is responsible of glucan remodeling at neutral-alkaline pH and is essential for morphogenesis and virulence. Due to the pH-regulated expression of PHR1, the phr1Δ phenotype is manifested at pH > 6 and its severity increases with the rise in pH. We exploited the pH-conditional nature of a PHR1 null mutant to analyze the impact of glucan remodeling on the hyphal transcriptional program and the role of chitin synthases in the hyphal wall stress (HWS) response.ResultsIn hyphal growth inducing conditions, phr1Δ germ tubes are defective in elongation, accumulate chitin, and constitutively activate the signaling pathways mediated by the MAP kinases Mkc1p, Cek1p and Hog1p. The transcriptional profiles revealed an increase of transcript levels for genes involved in cell wall formation (CHS2 and CHS8, CRH11, PGA23, orf19.750, RBR1, RBT4, ECM331, PGA6, PGA13), protein N-glycosylation and sorting in the ER (CWH8 and CHS7), signaling (CPP1, SSK2), ion transport (FLC2, YVC1), stress response and metabolism and a reduced expression of adhesins. A transient up-regulation of DNA replication genes associated with entry into S-phase occurred whereas cell-cycle regulating genes (PCL1, PCL2, CCN1, GIN4, DUN1, CDC28) were persistently up-regulated. To test the physiological relevance of altered CHS gene expression, phr1Δ chsxΔ (x = 2,3,8) mutant phenotypes were analyzed during the Y-H transition. PHR1 deletion was synthetic lethal with CHS3 loss on solid M199 medium-pH 7.5 and with CHS8 deletion on solid M199-pH 8. On Spider medium, PHR1 was synthetic lethal with CHS3 or CHS8 at pH 8.ConclusionsThe absence of Phr1p triggers an adaptive response aimed to reinforce the hyphal cell wall and restore homeostasis. Chs3p is essential in preserving phr1Δ cell integrity during the Y-H transition. Our findings also unveiled an unanticipated essential role of Chs8p during filamentation on solid media. These results highlight the flexibility of fungal cells in maintaining cell wall integrity and contribute to assessments of glucan remodeling as a target for therapy.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2853-5) contains supplementary material, which is available to authorized users.

Regulation of Neurospora crassa cell wall remodeling via the cot-1 pathway is mediated by gul-1.

Impairment of the Neurospora crassa Nuclear DBF2-related kinase-encoding gene cot-1 results in pleiotropic effects, including abnormally thick hyphal cell walls and septa. An increase in the transcript abundance of genes encoding chitin and glucan synthases and the chitinase gh18-5, but not the cell wall integrity pathway transcription factor rlm-1, accompany the phenotypic changes observed. Deletion of chs-5 or chs-7 in a cot-1 background results in a reduction of hyperbranching frequency characteristic of the cot-1 parent. gul-1 (a homologue of the yeast SSD1 gene) encodes a translational regulator and has been shown to partially suppress cot-1. We demonstrate that the high expression levels of the cell wall remodeling genes analyzed is curbed, and reaches near wild type levels, when gul-1 is inactivated. This is accompanied by morphological changes that include reduced cell wall thickness and restoration of normal chitin levels. We conclude that gul-1 is a mediator of cell wall remodeling within the cot-1 pathway.

Modifications to the composition of the hyphal outer layer of Aspergillus fumigatus modulates HUVEC proteins related to inflammatory and stress responses

Aspergillus fumigatus, the main etiologic agent causing invasive aspergillosis, can induce an inflammatory response and a prothrombotic phenotype upon contact with human umbilical vein endothelial cells (HUVECs). However, the fungal molecules involved in this endothelial response remain unknown. A. fumigatus hyphae produce an extracellular matrix composed of galactomannan, galactosaminogalactan and α-(1,3)-glucan. In this study, we investigated the consequences of UGM1 gene deletion in A. fumigatus, which produces a mutant with increased galactosaminogalactan production. The ∆ugm1 mutant exhibited an HUVEC-hyperadhesive phenotype and induced increased endothelial TNF-α secretion and tissue factor mRNA overexpression in this “semi-professional” immune host cell. Using a shotgun proteomics approach, we show that the A. fumigatus ∆ugm1 strain can modulate the levels of proteins in important endothelial pathways related to the inflammatory response mediated by TNF-α and to stress response pathways. Furthermore, a purified galactosaminogalactan fraction was also able to induce TNF-α secretion and the coincident HUVEC pathways regulated by the ∆ugm1 mutant, which overexpresses this component, as demonstrated by fluorescence microscopy. This work contributes new data regarding endothelial mechanisms in response to A. fumigatus infection. SignificanceInvasive aspergillosis is the main opportunistic fungal infection described in neutropenic hematologic patients. One important clinical aspect of this invasive fungal infection is vascular thrombosis, which could be related, at least in part, to the activation of endothelial cells, as shown in previous reports from our group. It is known that direct contact between the A. fumigatus hyphal cell wall and the HUVEC cell surface is necessary to induce an endothelial prothrombotic phenotype and secretion of pro-inflammatory cytokines, though the cell surface components of this angioinvasive fungus that trigger this endothelial response are unknown. The present work employs a discovery-driven proteomics approach to reveal the role of one important cell wall polysaccharide of A. fumigatus, galactosaminogalactan, in the HUVEC interaction and the consequent mechanisms of endothelial activation. This is the first report of the overall panel of proteins related to the HUVEC response to a specific and purified cell wall component of the angioinvasive fungus A. fumigatus.

The Sclerotinia sclerotiorum Slt2 mitogen-activated protein kinase ortholog, SMK3, is required for infection initiation but not lesion expansion

Mitogen-activated protein kinases (MAPKs) play a central role in transferring signals and regulating gene expression in response to extracellular stimuli. An ortholog of the Saccharomyces cerevisiae cell wall integrity MAPK was identified in the phytopathogenic fungus Sclerotinia sclerotiorum. Disruption of the S. sclerotiorum Smk3 gene severely reduced virulence on intact host plant leaves but not on leaves stripped of cuticle wax. This was attributed to alterations in hyphal apical dominance leading to the inability to aggregate and form infection cushions. The mutation also caused loss of the ability to produce sclerotia, increased aerial hyphae formation, and altered hyphal hydrophobicity and cell wall integrity. Mutants had slower radial expansion rates on solid media but more tolerance to elevated temperatures. Loss of the SMK3 cell wall integrity MAPK appears to have impaired the ability of S. sclerotiorum to sense its surrounding environment, leading to misregulation of a variety of functions. Many of the phenotypes were similar to those observed in S. sclerotiorum adenylate cyclase and SMK1 MAPK mutants, suggesting that these signaling pathways co-regulate aspects of fungal growth, physiology, and pathogenicity.

Biocontrol of Wheat Fusarium Crown and Root Rot by Trichoderma spp. and Evaluation of Their Cell Wall Degrading Enzymes Activities

Fourteen strains of Trichoderma spp. were isolated from Algerian desert soils and assessed for their antagonistic activity against Fusarium crown and root rot of wheat. Biocontrol efficiency of Trichoderma spp. was studied by in vitro and in vivo based bioassay against three pathogenic species: F. culmorum, F. graminearum and F. verticillioides. In vitro based bioassay (dual culture) results obtained with all Trichoderma spp. isolates showed significant decrease in colony diameter of Fusarium species compared to the control. The highest percentages of reduction in colony diameter were obtained with T. harzianum Thr.4 causing a growth reduction of 70.68%, 67.05 and 70.57% against F. culmorum, F. graminearum and F. verticillioides, respectively. All Trichoderma spp. isolates were able to overgrow and sporulate above F. culmorum colonies but no overgrowth was observed with F. graminearum and F. verticilliodes. The seed treatment by Trichoderma spp. isolates before sowing in a soil already infested by the pathogens led to a significant decrease of disease severity compared to the untreated control. The highest disease index decrease (&gt;70%) was obtained with two isolates of T. harzianum (Thr.4 and Thr.10) and T. viride Tv.6 against the three fungal pathogens. Lytic enzymes production by Trichoderma spp. isolates was tested in liquid cultures containing fungal cell walls of each pathogen as sole carbon source. Higher levels of protease and chitinase activities were induced by hyphal cell walls of F. graminearum than cell walls of F. verticillioides and F. culmorum. T. harzianum Thr.4 exhibited the highest enzyme activities with hyphal cell walls of F. graminearum and F. culmorum. However, in the medium amended with cell wall of F. verticillioides, maximal lytic activities were recorded for T. viride Tv.6.

Characterization of the Sclerotinia sclerotiorum cell wall proteome.

SummaryWe used a proteomic analysis to identify cell wall proteins released from Sclerotinia sclerotiorum hyphal and sclerotial cell walls via a trifluoromethanesulfonic acid (TFMS) digestion. Cell walls from hyphae grown in Vogel's glucose medium (a synthetic medium lacking plant materials), from hyphae grown in potato dextrose broth and from sclerotia produced on potato dextrose agar were used in the analysis. Under the conditions used, TFMS digests the glycosidic linkages in the cell walls to release intact cell wall proteins. The analysis identified 24 glycosylphosphatidylinositol (GPI)‐anchored cell wall proteins and 30 non‐GPI‐anchored cell wall proteins. We found that the cell walls contained an array of cell wall biosynthetic enzymes similar to those found in the cell walls of other fungi. When comparing the proteins in hyphal cell walls grown in potato dextrose broth with those in hyphal cell walls grown in the absence of plant material, it was found that a core group of cell wall biosynthetic proteins and some proteins associated with pathogenicity (secreted cellulases, pectin lyases, glucosidases and proteases) were expressed in both types of hyphae. The hyphae grown in potato dextrose broth contained a number of additional proteins (laccases, oxalate decarboxylase, peroxidase, polysaccharide deacetylase and several proteins unique to Sclerotinia and Botrytis) that might facilitate growth on a plant host. A comparison of the proteins in the sclerotial cell wall with the proteins in the hyphal cell wall demonstrated that sclerotia formation is not marked by a major shift in the composition of cell wall protein. We found that the S. sclerotiorum cell walls contained 11 cell wall proteins that were encoded only in Sclerotinia and Botrytis genomes.

Candida glabrata Binding to Candida albicans Hyphae Enables Its Development in Oropharyngeal Candidiasis.

Pathogenic mechanisms of Candida glabrata in oral candidiasis, especially because of its inability to form hyphae, are understudied. Since both Candida albicans and C. glabrata are frequently co-isolated in oropharyngeal candidiasis (OPC), we examined their co-adhesion in vitro and observed adhesion of C. glabrata only to C. albicans hyphae microscopically. Mice were infected sublingually with C. albicans or C. glabrata individually, or with both species concurrently, to study their ability to cause OPC. Infection with C. glabrata alone resulted in negligible infection of tongues; however, colonization by C. glabrata was increased by co-infection or a pre-established infection with C. albicans. Furthermore, C. glabrata required C. albicans for colonization of tongues, since decreasing C. albicans burden with fluconazole also reduced C. glabrata. C. albicans hyphal wall adhesins Als1 and Als3 were important for in vitro adhesion of C. glabrata and to establish OPC. C. glabrata cell wall protein coding genes EPA8, EPA19, AWP2, AWP7, and CAGL0F00181 were implicated in mediating adhesion to C. albicans hyphae and remarkably, their expression was induced by incubation with germinated C. albicans. Thus, we found a near essential requirement for the presence of C. albicans for both initial colonization and establishment of OPC infection by C. glabrata.

The endophytic symbiont Epichloë festucae establishes an epiphyllous net on the surface of Lolium perenne leaves by development of an expressorium, an appressorium-like leaf exit structure.

Summary Epichloë festucae forms a mutualistic symbiotic association with Lolium perenne. This biotrophic fungus systemically colonizes the intercellular spaces of aerial tissues to form an endophytic hyphal network. E. festucae also grows as an epiphyte, but the mechanism for leaf surface colonization is not known. Here we identify an appressorium‐like structure, which we call an expressorium that allows endophytic hyphae to penetrate the cuticle from the inside of the leaf to establish an epiphytic hyphal net on the surface of the leaf.We used a combination of scanning electron, transmission electron and confocal laser scanning microscopy to characterize this novel fungal structure and determine the composition of the hyphal cell wall using aniline blue and wheat germ agglutinin labelled with Alexafluor‐488.Expressoria differentiate immediately below the cuticle in the leaf blade and leaf sheath intercalary cell division zones where the hyphae grow by tip growth. Differentiation of this structure requires components of both the NoxA and NoxB NADPH oxidase complexes. Major remodelling of the hyphal cell wall occurs following exit from the leaf.These results establish that the symbiotic association of E. festucae with L. perenne involves an interconnected hyphal network of both endophytic and epiphytic hyphae.

Inhibition of Candida albicans biofilm formation and modulation of gene expression by probiotic cells and supernatant.

Oral candidiasis is a disease caused by opportunistic species of Candida that normally reside on human mucosal surfaces. The transition of Candida from budding yeast to filamentous hyphae allows for covalent attachment to oral epithelial cells, followed by biofilm formation, invasion and tissue damage. In this study, combinations of Lactobacillus plantarum SD5870, Lactobacillus helveticus CBS N116411 and Streptococcus salivarius DSM 14685 were assessed for their ability to inhibit the formation of and disrupt Candida albicans biofilms. Co-incubation with probiotic supernatants under hyphae-inducing conditions reduced C. albicans biofilm formation by >75 % in all treatment groups. Likewise, combinations of live probiotics reduced biofilm formation of C. albicans by >67 %. When live probiotics or their supernatants were overlaid on preformed C. albicans biofilms, biofilm size was reduced by >63 and >65 % respectively. Quantitative real-time PCR results indicated that the combined supernatants of SD5870 and CBS N116411 significantly reduced the expression of several C. albicans genes involved in the yeast-hyphae transition: ALS3 (adhesin/invasin) by 70 % (P < 0.0001), EFG1 (hyphae-specific gene activator) by 47 % (P = 0.0061), SAP5 (secreted protease) by 49 % (P < 0.0001) and HWP1 (hyphal wall protein critical to biofilm formation) by >99 % (P < 0.0001). These findings suggest the combination of L. plantarum SD5870, L. helveticus CBS N116411 and S. salivarius DSM 14685 is effective at both preventing the formation of and removing preformed C. albicans biofilms. Our novel results point to the downregulation of several Candida genes critical to the yeast-hyphae transition, biofilm formation, tissue invasion and cellular damage.

Nanoscopic Cell Wall Architecture of an Immunogenic Ligand in Candida Albicans during Antifungal Drug Treatment

Candida albicans is a common cause of high-mortality disseminated fungal infections. Morphotype switching between yeasts and filamentous hyphae is important for virulence. Innate immune cells use C-type lectins to recognize Candida cell wall polysaccharides. In particular, Dectin-1 recognizes fungal cell wall beta-glucan, leading to phagocytosis and pro-inflammatory responses. However, C. albicans partially evades innate immune recognition by restricting glucan surface exposure. Treatment with the antifungal caspofungin increases surface exposure of beta-glucan and leukocyte phagocytosis. Our objective was to better understand the nanoscale structure of glucan exposures that C. albicans presents to leukocytes. We probed exposed glucan with AF647-soluble Dectin-1 ectodomain and used direct Stochastic Optical Reconstruction Microscopy (dSTORM) to quantify glucan exposure on C. albicans yeast and hyphal cell walls, both in the resting state and after treatment with caspofungin. Strikingly, the vast majority of glucan exposure sites were consistent with single receptor binding sites in all cases. In resting yeasts, the multiple-probe exposure sites were few and small. After caspofungin treatment, multi-probe exposures were larger and surface density of glucan exposures was increased. Untreated C. albicans hyphae had a higher glucan exposure surface density than yeasts. Lateral hyphal cell walls imaged by dSTORM underwent little quantitative or qualitative change in glucan exposure geometry after drug treatment, but confocal imaging revealed higher glucan exposure after caspofungin treatment at the actively growing and remodeling cell walls of hyphal tips. Similarly, in budding yeasts, we found consistently higher glucan exposure in the actively remodeling cell wall of daughter cells. These findings enable future mechanistic investigation of the contribution of glucan nanopatterning to C. albicans evasion of innate immune recognition via Dectin-1 and the use of nanopatterning of glucan for therapeutic interventions in fungal infectious disease.

Design and studies of multiple mechanism of anti-Candida activity of a new potent Trp-rich peptide dendrimers

PurposeEight peptide dendrimers were designed as structural mimics of natural cationic amphiphilic peptides with antifungal activity and evaluated for their anti-Candida potential against the wild type strains and mutants. MethodsDendrimer 14 containing four Trp residues and dodecyl tail and a slightly smaller dendrimer 9 decorated with four N-methylated Trp that displayed 100 and 99.7% of growth inhibition at 16 μg/mL respectively, were selected for evaluation against the Candida albicans mutants with disabled biosynthesis of aspartic proteases responsible for host tissue colonization and morphogenesis during biofilm formation (sessile model). Flow cytometry method was employed to detect apoptotic cells with membrane alterations (phosphatidylserine translocation), and differentiation of apoptotic from necrotic cells was also performed. Simultaneous staining of cell surface phosphatidylserine with Annexin-V-Fluorescein and necrotic cells with propidium iodide was conducted. Results14 at 16 μg/mL caused C. albicans cells to undergo cellular apoptosis but its increasing concentrations induced necrosis. 14 influenced C. albicans biofilm viability as well as hyphal and cell wall morphology. Confocal microscopy and cell wall staining with calcofluor white revealed that in epithelial model the cell surface structure was perturbed at MIC of peptide dendrimer. It appears that tryptophan or 1-methyltryptophan groups displayed at the surface and positive charges hidden in the dendrimer tree along with hydrocarbon tail located at C-terminus are important for the anti-Candida activity since dendrimers containing tryptamine at C-terminus showed only a moderate activity. ConclusionsOur results suggest that membranolytic dendrimer 14, targeting cellular apoptotic pathway and impairing the cell wall formation in mature biofilm, may be a potential multifunctional antifungal lead compound for the control of C. albicans infections.

Response of the fungus Pseudocercospora griseola f. mesoamericana to Tricyclazole

Pseudocercospora griseola, an anamorph of Mycosphaerella, causes Angular Leaf Spot (ALS). The mycelia and conidia from P. griseola are coloured due to the synthesis of 1,8 dihydroxynaphthalene (DHN)-melanin. The aim of this work was to identify in P. griseola f. mesoamericana isolate T4, a highly pigmented fungus, intermediary compounds as a result of the inhibition of melanin synthesis by tricyclazole, and to analyze at the structural level the localization of these dark pigments. The main metabolites were analyzed using ultraviolet matrix-assisted laser desorption-ionization mass spectrometry (UV–MALDI MS). Tricyclazole affected P. griseola f. mesoamericana in several different ways. The most evident effect was the reduction of melanin synthesis, and therefore diffusible shunt products were found and identified. Flaviolin was the main intermediate metabolite found in cultures supplemented with tricyclazole. This inhibitor, which affected pigmentation and the cell wall structure of mycelium, revealed macroscopically by the reduction in growth, decreased the stratification and deposition of melanin in the hyphal wall. These results suggest a possible role of tricyclazole to control ALS.

Identification and conservation of a Neolithic polypore

Abstract An archaeological waterlogged polypore was found in the Final Jomon period (circa 2600 years ago) at the Neolithic site of Kitashirakawa in Kyoto, Japan. The fragile polypore was treated with the feather-keratin method that we developed for the preservation of wet organic archaeological materials. The method reinforced the polypore against the collapse caused by air-drying. The anatomical features of the polypore were observed under light microscopy and the host wood present at the edge of the basidiocarp was identified as Morus sp. on the basis of anatomical observations. Eventually, the polypore was identified as Inonotus sanghuang . Macroscopic and microscopic observations were carried out in order to evaluate the effectiveness of the keratin treatment on the polypore tissue. The keratin consolidated to the hyphal cell walls, where it stabilized the thick walled fungal mycelium that makes up the polypore fruiting body.

Mannoproteins from yeast and hyphal form of Candida albicans considerably differ in mannan and protein content

Significant differences in carbohydrate composition of mannoproteins obtained from yeast and hyphal cell walls of Candida albicans (serotypes A and B) were found. Yeast mannoproteins from both serotypes consisted up to 46% of mannan while the same parts from hyphal cells contained only about 14% of mannan. Another difference was in protein content, 47–53% for yeasts, 3–4.5% for hyphae, respectively. Moreover, HPLC profiles of yeast mannoproteins were more complex compared to those of hyphal form. Subsequently, mannans were prepared from yeast and hyphal mannoproteins using cetavlon fractionation. Mannans from both yeast serotypes contained higher amounts of mannose (91.4% serotype A; 92.8% serotype B) than mannans from hyphae (66.4% serotype A; 76.3% serotype B). Unlike mannans from serotype B, mannans from serotype A contained β-(1→2)-linked mannopyranosyl units in acid-stable moiety. Further, hyphal mannans were less branched than yeast mannans. The shift from yeast to hyphal form probably led to simplification of mannan structure.